Morphological and molecular characterization of Strongyloides ophidiae (Nematoda, Strongyloididae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Spirorchiids (Digenea: Spirorchiidae) infecting a Hawksbill sea turtle Eretmochelys imbricata (Linnaeus 1758) from Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Relationship of intestinal histology with the resistance to Trichostrongylus colubriformis infection in three breeds of sheep

Avaliaram-se a associação entre o número de células inflamatórias no intestino delgado e a resistência à infecção por Trichostrongylus colubriformis em ovinos de três raças (Santa Inês, Suffolk e Ile de France), naturalmente infectados. Mastócitos, eosinófilos e leucócitos globulares foram quantificados na mucosa intestinal. A concentração de histamina foi estimada em amostras teciduais do intestino, bem como foi determinado o comprimento de machos e fêmeas de T. colubriformis. A resposta celular foi similar na mucosa intestinal das três raças ovinas (P>0,05). Houve grande variação entre os ovinos em relação aos resultados parasitológicos e celulares, mesmo nos animais de mesma raça. em geral, os animais que apresentaram número menor de células inflamatórias tiveram cargas parasitárias maiores, contagens de ovos por grama de fezes mais altas e exemplares de T. colubriformis maiores. Os resultados indicaram que mastócitos, eosinófilos e leucócitos globulares prejudicaram o estabelecimento, o desenvolvimento e a sobrevivência dos parasitas.

Faecal examination and PCR to detect Strongyloides venezuelensis in experimentally infected Lewis rats

More sensitive methodologies are necessary to improve strongyloidiasis diagnosis. This study compared the sensitivities of the McMaster modified technique and polymerase chain reaction (PCR) assays, both performed in faecal samples. Lewis rats were subcutaneously infected with 4,000, 400 or 40 infective third-stage larvae, considered as high, moderate or low infection, respectively. Seven days later, they were euthanized to count adult nematodes recovered from the small intestine. Stool samples were used to count the number of eggs per gram (EPG) of faeces and to detect parasite DNA by PCR performed with a species and a genus primer pair. The sensitivity of these assays depended upon parasite burden and the primer specificity. All assays presented 100% sensitivity at the highest parasite load. In the moderate infection, EPG and PCR with the genus primer maintained 100% specificity, whereas PCR sensitivity with the species primer decreased to 77.7%. In low infection, the sensitivity was 60% for EPG, 0% for PCR with the species primer and 90% for PCR done with the genus primer. Together, these results suggest that PCR with a genus primer can be a very sensitive methodology to detect Strongyloides venezuelensisin faeces of Lewis rats infected with very low parasite burden.

Strongyloides venezuelensis infection susceptibility of seven inbred strains of mice

Foi investigada a susceptibilidade de sete linhagens isogênicas de camundongos à infecção experimental, primária e secundária, por Strongyloides venezuelensis a fim de servir de base para estudos genéticos sobre a resistência. Foram utilizados 12 camundongos machos, com seis semanas de idade, das seguintes linhagens isogênicas: A/J, BALB/c, CBA/J, C3H/Hepos, C57BL/6, DBA/2 e NIH. Os animais foram inoculados, via sub-cutânea, com 2000 larvas infectantes. As contagens médias (± desvio padrão) de parasitas no intestino delgado dos camundongos seis dias após a infecção, em ordem crescente, foram: 28 (± 19) na linhagem NIH; 647 (± 228) na BALB/c; 709 (± 425) na DBA/2; 731 (± 151) na C3H/Hepos, 801 (± 174) na CBA/J; 1024 (± 267) na C57BL/6 e 1313 (± 483) na A/J. Os camundongos C57BL/6 apresentaram as mais elevadas contagens de ovos de S. venezuelensis por grama de fezes (OPG) e os NIH, as mais baixas. Não foram detectados ovos nos exames de fezes e não foram encontrados parasitas no intestino delgado dos animais re-infectados 14 dias após a infecção primária. A linhagem NIH apresentou elevada resistência contra as infecções primárias por S. venezuelensis. Entre as outras seis linhagens, uma das mais susceptíveis foi a linhagem C57BL/6.

Immunological responses and cytokine gene expression analysis to Cooperia punctata infections in resistant and susceptible Nelore cattle

Cellular and humoral immune response, as well as cytokine gene expression, was assessed in Nelore cattle with different degrees of resistance to Cooperia punctata natural infection. One hundred cattle (male, weaned, 11-12 months old), kept together on pasture, were evaluated. Faecal and blood samples were collected for parasitological and immunological assays. Based on nematode faecal egg counts (FEC) and worm burden, the seven most resistant and the eight most susceptible animals were selected. Tissue samples of the small intestine were collected for histological quantification of inflammatory cells and analysis of cytokine gene expression (IL-2, IL-4, IL-8, IL-1 2p35, IL-13, TNF-alpha, IFN-gamma, MCP-1, MCP-2, and MUC- 1) using real-time RT-PCR. Mucus samples were also collected for IgA levels determination. Serum IgG1 mean levels against C. punctata antigens were higher in the resistant group, but significant differences between groups were only observed 14 days after the beginning of the experiment against infective larvae (1-3) and 14 and 84 days against adult antigens. The resistant group also presented higher IgA levels against C. punctata (L3 and adult) antigens with significant difference 14 days after the beginning of the trial (P < 0.05). In the small-intestine mucosa, levels of IgA anti-L3 and anti-adult C. punctata were higher in the resistant group, compared with the susceptible group (P < 0.05). Gene expression of both T(H)2 cytokines (IL-4 and IL-13) in the resistant group and T(H)1 cytokines (IL-2, IL-1 2p35, IFN-gamma and MCP-1) in the susceptible group was up-regulated. Such results suggested that immune response to C. punctata was probably mediated by TH2 cytokines in the resistant group and by T(H)1 cytokines in the susceptible group. (C) 2008 Elsevier B.V. All rights reserved.

Identification and characterization of excretory/secretory products of Giardia duodenalis trophozoites

One of the major questions concerning Giardia is the understanding of pathophysiological processes associated with small intestine abnormalities. There are evidences that Giardia trophozoites contain and/or release proteolytic enzymes that may be implicated in the host intestinal epithelium. The present investigation was undertaken to examine the protease activity in excretory/secretory (E/S) products of Giardia duodenalis trophozoites of an axenic Brazilian strain (BTU-11) and the reference strain Portland 1 (P1). E/S products from trophozoites of each strain in conditioned medium were tested with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for the protein profiles, and the protease activity was analyzed using substrate-impregnated SDS-PAGE (gelatin and collagen) and hemoglobin assay. The proteases characterization was based on inhibition assays including synthetic inhibitors. Electrophoresis analysis of E/S products revealed a banding pattern composed by few bands (4 to 6 bands) in the migration region of 123 to 28 kDa. Proteolytic products were detected in the conditioned medium by trophozoites of both assayed strains. In the gels containing copolymerized gelatin and collagen, E/S products promoted substrate degradation and the most evident proteolysis zones were distributed in the migration regions of 77 to 18 kDa and 145 to 18 kDa, respectively, in the patterns of gelatinolytic and collagenolytic activities. Degradation of hemoglobin was also observed, and the pattern of hydrolysis was similar in both E/S products assayed. Inhibitor assays showed that the main proteolytic activity in both E/S products is due to cysteine proteases, although the presence of serine proteases was also indicated. Degradation of substrates including collagen and hemoglobin could lead us to speculate different functions of Giardia excreted/secreted proteases in vivo, but to confirm this possibility and to elucidate its implication on host-parasite interactions, further experiments applying protocols for the purification of proteases are necessary. Even so, our observations are relevant and hold the perspective for the understanding about protease activity in Giardia trophozoites of axenic strain isolated in an endemic area.

Immune responses in sheep naturally infected with Oestrus ovis (Diptera: Oestridae) and gastrointestinal nematodes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Paracellular Absorption: A Bat Breaks the Mammal Paradigm

Bats tend to have less intestinal tissue than comparably sized nonflying mammals. The corresponding reduction in intestinal volume and hence mass of digesta carried is advantageous because the costs of flight increase with load carried and because take-off and maneuverability are diminished at heavier masses. Water soluble compounds, such as glucose and amino acids, are absorbed in the small intestine mainly via two pathways, the transporter-mediated transcellular and the passive, paracellular pathways. Using the microchiropteran bat Artibeus literatus (mean mass 80.6 +/- 3.7 g), we tested the predictions that absorption of water-soluble compounds that are not actively transported would be extensive as a compensatory mechanism for relatively less intestinal tissue, and would decline with increasing molecular mass in accord with sieve-like paracellular absorption. Using a standard pharmacokinetic technique, we fed, or injected intraperitonealy the metabolically inert carbohydrates L-rhamnose (molecular mass = 164 Da) and cellobiose (molecular mass = 342 Da) which are absorbed only by paracellular transport, and 3-O-methyl-D-glucose (3OMD-glucose) which is absorbed via both mediated (active) and paracellular transport. As predicted, the bioavailability of paracellular probes declined with increasing molecular mass (rhamnose, 90 +/- 11%; cellobiose, 10 +/- 3%, n = 8) and was significantly higher in bats than has been reported for laboratory rats and other mammals. In addition, absorption of 3OMD-glucose was high (96 +/- 11%). We estimated that the bats rely on passive, paracellular absorption for more than 70% of their total glucose absorption, much more than in non-flying mammals. Although possibly compensating for less intestinal tissue, a high intestinal permeability that permits passive absorption might be less selective than a carrier-mediated system for nutrient absorption and might permit toxins to be absorbed from plant and animal material in the intestinal lumen.

Ecological aspects of helminth fauna of Magellanic penguins, Spheniscus magellanicus (aves: Spheniscidae), from the Northern Coast of the State of São Paulo, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Maternal protein restriction affects gene expression and enzyme activity of intestinal disaccharidases in adult rat offspring

This study investigated the consequences of intrauterine protein restriction on the gastrointestinal tract and particularly on the gene expression and activity of intestinal disaccharidases in the adult offspring. Wistar rat dams were fed isocaloric diets containing 6% protein (restricted, n = 8) or 17% protein (control, n = 8) throughout gestation. Male offspring (n = 5-8 in each group) were evaluated at 3 or 16 weeks of age. Maternal protein restriction during pregnancy produced offspring with growth restriction from birth (5.7 ± 0.1 vs 6.3 ± 0.1 g; mean ± SE) to weaning (42.4 ± 1.3 vs 49.1 ± 1.6 g), although at 16 weeks of age their body weight was similar to control (421.7 ± 8.9 and 428.5 ± 8.5 g). Maternal protein restriction also increased lactase activity in the proximal (0.23 ± 0.02 vs 0.15 ± 0.02), medial (0.30 ± 0.06 vs 0.14 ± 0.01) and distal (0.43 ± 0.07 vs 0.07 ± 0.02 U·g-1·min-1) small intestine, and mRNA lactase abundance in the proximal intestine (7.96 ± 1.11 vs 2.38 ± 0.47 relative units) of 3-week-old offspring rats. In addition, maternal protein restriction increased sucrase activity (1.20 ± 0.02 vs 0.91 ± 0.02 U·g-1·min-1) and sucrase mRNA abundance (4.48 ± 0.51 vs 1.95 ± 0.17 relative units) in the duodenum of 16-week-old rats. In conclusion, the present study shows for the first time that intrauterine protein restriction affects gene expression of intestinal enzymes in offspring.

Postweaning growth of the non-carcass components of Texel crossbred lambs

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Effect of the levels of calcium and particle size of limestone on laying hens

O experimento foi realizado para avaliar o efeito dos níveis de cálcio (Ca) e da granulometria do calcário (GC) para poedeiras comerciais. Foram utilizadas 216 poedeiras da linhagem Dekalb White de 25 a 49 semanas de idade em um delineamento inteiramente casualizado, em esquema fatorial 3 × 2, composto de três níveis de cálcio (3,92; 4,02 e 4,12%) e duas granulometrias do calcário (fina - 0,60 mm; e grossa - 1,00 mm), que resultaram em seis tratamentos, com seis repetições de seis aves. Não houve efeito significativo dos níveis de cálcio e da granulometria do calcário nem da interação nível de cálcio × granulometria do calcário para o consumo de ração, o peso dos ovos, a gravidade específica, a espessura da casca, a porcentagem de casca, a unidade Haugh, a digestibilidade de cálcio e fósforo e a porcentagem do trato digestório. A produção, massa, conversão por massa e conversão por dúzia de ovos tiveram seus melhores resultados com 4,12% de cálcio na dieta. A porcentagem de moela e o consumo de ração observado às 15 h melhoraram com o aumento na granulometria do calcário de 0,60 para 1,00 mm. Houve efeito da interação entre nível de cálcio e granulometria do calcário para a resistência à quebra da tíbia, a densidade óssea, o comprimento do intestino delgado e o consumo de ração às 6, 9, 12, 18 e 21 h, respectivamente. Recomenda-se a adição de 4,12% de cálcio e calcário na granulometria grossa (1,00 mm) para melhor desempenho de poedeiras comerciais. Mais estudos são necessários para elucidar os mecanismos envolvidos na alteração do comportamento alimentar das poedeiras em resposta às mudanças nos níveis de cálcio e na granulometria do calcário das rações.

Rendimento dos componentes não-carcaça de cordeiros alimentados com silagem de milho ou cana-de-açúcar e dois níveis de concentrado

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Jejum Pré-abate em Frangos de Corte

As condições ideais dos frangos de corte no momento do abate devem ser conhecidas a fim de possibilitar a produção de carne de excelente qualidade, uma vez que diversos fatores pré e pós-abate estão envolvidos na qualidade final. em condições normais de abate e processamento, a retirada de ração é feita de 6 a 8 horas antes da apanha das aves, resultando em um período total de jejum de 8 a 12 horas antes do abate, para esvaziar o intestino e com isso minimizar a contaminação no abatedouro. A escalda, depena e evisceração são pontos importantes de contaminação cruzada no abatedouro devido à grande quantidade de microorganismos aderidos às penas, pele e patas das aves e ao rompimento das vísceras durante a evisceração. Entretanto, a desidratação da carcaça começa imediatamente após o início do jejum. Períodos prolongados de jejum podem afetar o pH das diversas partes do intestino, aumentando a presença de Salmonella e outros microorganismos patogênicos. Além disso, determinam uma maior contaminação pela bile, e são, subjetivamente, associados à fragilidade dos intestinos durante a evisceração mecânica. Portanto, os esquemas de processamento devem ser estabelecidos levando-se em conta a integridade e o esvaziamento do intestino e da vesícula biliar, bem como a desidratação e os seus efeitos sobre o bem estar das aves, contaminação da carcaça e qualidade da carne. Como alguns efeitos do jejum ainda não são bem conhecidos, sugerem-se pesquisas nas seguintes áreas: definir o tempo ótimo de jejum para atender o bem estar das aves, minimizar a contaminação e otimizar os parâmetros de qualidade de carcaça; estudar os efeitos de períodos prolongados de jejum sobre o pH e a colonização do papo, pró-ventrículo, moela, intestino delgado, intestino grosso e cecos por enterobactérias, como Salmonella, por exemplo; efeito do jejum sobre o tamanho e cor do fígado. O resultado esperado é um aumento na qualidade final dos produtos aliado a uma redução nas perdas e no custo de produção.